Neural crest cells are a population of migratory, multipotent cells that differentiate into many diverse vertebrate lineages, including the craniofacial skeleton, peripheral nervous system, and melanocytes. Deficiencies in neural crest development lead to a variety of birth defects, including craniofacial clefts and some congenital heart defects. Folate supplements reduce the incidence of these birth defects, but the mechanism of this effect is unknown. One possibility is that folates rescue methylation. Although methylation has never been implicated in neural crest development, my preliminary data show that methylation is required for neural crest migration. Based on the expression of the methyltransferase Ezh2 in the cytoplasm of migratory neural crest cells, I postulate that non-histone protein methylation controls the migratory properties of neural crest cells. This proposal seeks to test this hypothesis and to identify and characterize the methyltransferases that mediate methylation in the neural crest through three specific aims: (1) The spatial and temporal localization of three candidate methyltransferases will be determined by immunofluorescence. (2) Antisense morpholino oligonucleotide-mediated protein ablation will be used to assay the requirement for each candidate methyltransferases during neural crest development. (3) The specific requirement for the methyltransferase Ezh2 in the cytoplasm and the nucleus will be assessed using dominant negative constructs. Once the requirement for particular methyltransferases is established in the neural crest, we can begin to look at the effects of folate-deficiency on methylation, and elucidate the mechanism by which folate supplementation reduces birth defects. This work will also provide insight into the molecular mechanisms of cancer metastasis. PUBLIC HEALTH RELEVANCE: Neural crest cells are cells that move through the developing embryo to form parts of the nervous system, face, heart and skin. The goal of this research is to determine how neural crest cells are able to move throughout the body. This process is similar to cancer and will help us to understand cancer progression, and will also provide us with a better understanding of how defects in neural crest cell movement can lead to birth defects.